Description: (Adapted from the application). The project will continue the ongoing efforts in studying cell specific expression and the structure and regulation of two hexokinase gene family members, glucokinase (GK) and hexokinase II (HKII). There are three major specific aims. The experiments proposed under the first specific aim will identify the regulatory elements in the mouse GK gene necessary for transcription in liver. We will identify DNase I HS-sites flanking the downstream promoter in the gene, perform comparative DNA sequence analysis of mammalian GK genes, identify liver-specific DNA-protein interactions, and test the function of a 83 kb fragment of the gene in transgenic mice. The experiments proposed under the second specific aim, are to insert a LacZ reporter gene into the first exon of the HKII gene and liver-specific first exon of the GK gene using gene targeting methods. The object of these experiments is to produce mice which will be useful in answering questions related to both the function and cell- specific expression of GK and HKII. The experiments proposed under the third specific aim will analyze the factors and elements that determine the cell-specific expression of the islet GK isoform. Efforts will be directed towards determining the role of a pancreatic-duodenal homeobox gene (PDX-l) on expression of GK in the beta cell to the cloning of proteins that bind to the Pal elements in the upstream promoter, and to the identification and functional characterization of silencer elements that may contribute to the cell-specific expression of the islet GK isoform.